Nucleus Pulposus of the Intervertebral Disc: The Effect of Environmental Factors on Metabolic Activity

Lower back pain is a major socioeconomic concern in developed nations. Moreover, it is a condition which a majority of people will experience an episode within their lifetime, with a portion becoming chronically afflicted. Degeneration of the intervertebral disc (IVD) is believed to play a critical role in initiating lower back pain. Poor nutrient supplies are implicated as a one of the root causes of this degeneration and much effort has been spent to better elucidate the behavior of IVD cells. The IVD is the largest avascular structure within the body, relying on diffusion to migrate nutrients in and clear wastes out. Due to this limitation, nutrients concentrations are low within the center of the disc and waste products also accumulate in high concentrations. For most cell niches, this is considered a harsh environment, but completely normal for the cells of the IVD. Understanding how these cells behave under these environmental conditions may elucidate what nutritional and environmental factors lead to aberrant IVD cell behavior. This dissertation will explore the relationship between nutrient conditions and the metabolic adaptations of the IVD cells. Additionally, a custom bioreactor was constructed in order to study the complex IVD organ as a whole under varied nutritional conditions. Glucose consumption rate and gene expression of nucleus pulposus (NP) cells was investigated in an agarose gel system over prolonged culture periods with varied oxygen tension and glucose concentration treatments. Glucose consumption rate was found to decrease with increasing oxygen tension and over time but not with changing glucose concentration. Catabolic gene expression increased for all groups over time, with inhibitors of catabolism following suit. Collagen Type I increased in expression with time for the high oxygen tensions while other anabolic genes did not show any consistent trends over the culture period. The optimal reference genes were evaluated and the genes RPL4 and YWHAZ were found to be more stable than the commonly used 18s and GAPDH. Next the glucose consumption rate was modeled using a more sophisticated method, the Michalis-Menten kinetic model, allowing for increased fidelity in computational modeling and optimizing culture conditions. No differences between oxygen tension were found in the GCR. Gene expression was again analyzed but the limited culture time did not allow for differences to be seen between oxygen tensions. A correlational analysis revealed several targets for further genetic studies. A custom-built bioreactor was developed and validated to load whole IVD under simulated physiological loading conditions. This reactor was then used to test whole IVDs under free-swelling and dynamic loading conditions. Tissues were compared for their gross composition as well as gene expression differences. No differences were seen between loaded and free-swelling discs, and both tended toward a catabolic state when examining gene expression. Gross changes in tissue composition were seen briefly in hydroxyproline content before returning to baseline. The contents of this dissertation greatly enhance the knowledge of IVD NP cell metabolism. Given that these cells play a vital role in disc degeneration and will be critical in regenerative strategies to treat said condition, understanding the behavior of these critical cells is of paramount importanc

Effects of Oxygen Concentration and Culture Time on Porcine Nucleus Pulposus Cell Metabolism: An in vitro Study

Low back pain is a common ailment that affects millions of individuals each year and is linked to degeneration of the intervertebral discs in the spine. Intervertebral disc degeneration is known to result from an imbalance in anabolic and catabolic activity by disc cells. Due to the avascular nature of the intervertebral disc, oxygen deficiency may occur in the central nucleus pulposus (NP). The resulting hypoxia affects matrix regulation and energy metabolism of disc cells, although the mechanisms are not fully understood. This study investigates in vitro glucose consumption and gene expression by NP cells over time under varying oxygen tensions. Notochordal porcine NP cells were cultured in agarose discs at 21, 5, or 1% oxygen tension for 1, 5, or 10 days. The expression of 10 key matrix genes, as well as Brachyury (T), by NP cells was analyzed using RT-PCR. Glucose consumption was measured using a two-point method. Results show that culture time and oxygen tension significantly affect glucose consumption rates by porcine NP cells. There were also significant changes in T expression based on oxygen level and culture time. The 1% oxygen tension had a significantly higher T expression on day 10 than the other two groups, which may indicate a better maintenance of the notochordal phenotype. MMP 1 and 13 expression increased over time for all groups, while only the 5% group showed an increase over time for MMP 3. TIMP expression followed the direction of MMPs but to a lesser magnitude. Five percent and twenty-one percent oxygen tensions led to decreases in anabolic gene expression while 1% led to increases. Oxygen concentration and culture time significantly impacted glucose consumption rate and the gene expression of matrix regulatory genes with hypoxic conditions most accurately maintaining the proper NP phenotype. This information is valuable not only for understanding disc pathophysiology, but also for harnessing the potential of notochordal NP cells in therapeutic applications

Aerogel Waveplates

Optical transmission measurements were made on 98% porosity silica aerogel samples under various degrees of uniaxial strain. Uniaxially compressed aerogels exhibit large birefringence, proportional to the amount of compression, up to the 15% strain studied. The birefringence is mostly reversible and reproducible through multiple compression-decompression cycles. Our study demonstrates that uniaxially strained high porosity aerogels can be used as tunable waveplates in a broad spectral range.Comment: 7 pages, 4 figures, submitted to Optics Expres

Drucker\u27s Insights on Market Orientation and Innovation: Implications for Emerging Areas in High-Technology Marketing

In 1954, Drucker boldly declared that organizations have only two basic functions, marketing and innovation. While true for any organization, this insight is particularly pertinent for technology-based businesses. The complicated environment surrounding high-tech companies creates a great need for sophisticated marketing, yet these companies continue to have under-developed competencies in marketing and in understanding customer needs. In its first two sections, this essay explores Drucker’s insights with respect to two particularly salient issues for high-tech companies: developing and implementing a market orientation, and sustained break-through innovations. We review Drucker’s insights and synthesize them with the scholarly research on these issues. In the third section, we discuss three emerging areas in high-tech marketing where academics and managers could build on Drucker’s insight to guide future research and practice: market-driving, customer co-creation, and corporate social responsibility. The illustrative examples provided by these emerging areas highlight that even today, Drucker’s writings continue to offer remarkable guidance to scholars and managers who are willing to take the time to reflect, understand, and incorporate these insights in the unique context of high-tech industries

Effect of Static Compressive Strain, Anisotropy, and Tissue Region on the Diffusion of Glucose in Meniscus Fibrocartilage

Osteoarthritis (OA) is a significant socio-economic concern, affecting millions of individuals each year. Degeneration of the meniscus of the knee is often associated with OA, yet the relationship between the two is not well understood. As a nearly avascular tissue, the meniscus must rely on diffusive transport for nutritional supply to cells. Therefore, quantifying structure–function relations for transport properties in meniscus fibrocartilage is an important task. The purpose of the present study was to determine how mechanical loading, tissue anisotropy, and tissue region affect glucose diffusion in meniscus fibrocartilage. A one-dimensional (1D) diffusion experiment was used to measure the diffusion coefficient of glucose in porcine meniscus tissues. Results show that glucose diffusion is strain-dependent, decreasing significantly with increased levels of compression. It was also determined that glucose diffusion in meniscus tissues is anisotropic, with the diffusion coefficient in the circumferential direction being significantly higher than that in the axial direction. Finally, the effect of tissue region was not statistically significant, comparing axial diffusion in the central and horn regions of the tissue. This study is important for better understanding the transport and nutrition-related mechanisms of meniscal degeneration and related OA in the knee

Conditional deletion of ferritin h in mice reduces B and T lymphocyte populations.

The immune system and iron availability are intimately linked as appropriate iron supply is needed for cell proliferation, while excess iron, as observed in hemochromatosis, may reduce subsets of lymphocytes. We have tested the effects of a ferritin H gene deletion on lymphocytes. Mx-Cre mediated conditional deletion of ferritin H in bone marrow reduced the number of mature B cells and peripheral T cells in all lymphoid organs. FACS analysis showed an increase in the labile iron pool, enhanced reactive oxygen species formation and mitochondrial depolarization. The findings were confirmed by a B-cell specific deletion using Fth(lox/lox) ; CD19-Cre mice. Mature B cells were strongly under-represented in bone marrow and spleen of the deleted mice, whereas pre-B and immature B cells were not affected. Bone marrow B cells showed increased proliferation as judged by the number of cells in S and G2/M phase as well as BrdU incorporation. Upon in vitro culture with B-cell activating factor of the tumor necrosis factor family (BAFF), ferritin H-deleted spleen B cells showed lower survival rates than wild type cells. This was partially reversed with iron-chelator deferiprone. The loss of T cells was also confirmed by a T cell-specific deletion in Fth(lox/lox) ;CD4-Cre mice. Our data show that ferritin H is required for B and T cell survival by actively reducing the labile iron pool. They further suggest that natural B and T cell maturation is influenced by intracellular iron levels and possibly deregulated in iron excess or deprivation

Market Orientation and the New Product Paradox.

The extant literature shows that the strength of the market orientation–performance relationship decays as the terminal measure of performance shifts from new product success to profitability to market share. As Day (1999) concluded, a broader nomological inquiry is needed to more fully understand the nature and limits of market orientation\u27s effects. This suggests that a broader nomological inquiry is needed to fully understand the nature and limits of market orientation\u27s effects. Utilizing a national sample of marketing executives, the present study\u27s purpose is to build a fuller understanding of the effects of market orientation on firm performance. Its structural equations model includes measures of new product success, profitability, and market share. The research reinforces a strong positive relationship between market orientation and new product success. The expanded nomological network under study, however, implies barriers to market orientation\u27s effectiveness. First, market-orientation-inspired increases in the priority firms place on “breakthrough” learning without commensurate increases in the priority placed on “breakthrough” innovation capabilities can boomerang and negatively impact new product success. Second, market-orientation-inspired new product development programs that are unable to increase market share can negatively impact profitability. These gatekeepers to the success of market orientation underscore the need for firms to coordinate a strong market orientation with resources and capabilities that increase the effectiveness of the marketing function. Without such coordination, the positive effect of market orientation on new product success may be limited to incremental innovations, and the overall effect of successful new products on profitability may be limited

Neuron Specific Rab4 Effector GRASP-1 Coordinates Membrane Specialization and Maturation of Recycling Endosomes

The endosomal pathway in neuronal dendrites is essential for membrane receptor trafficking and proper synaptic function and plasticity. However, the molecular mechanisms that organize specific endocytic trafficking routes are poorly understood. Here, we identify GRIP-associated protein-1 (GRASP-1) as a neuron-specific effector of Rab4 and key component of the molecular machinery that coordinates recycling endosome maturation in dendrites. We show that GRASP-1 is necessary for AMPA receptor recycling, maintenance of spine morphology, and synaptic plasticity. At the molecular level, GRASP-1 segregates Rab4 from EEA1/Neep21/Rab5-positive early endosomal membranes and coordinates the coupling to Rab11-labelled recycling endosomes by interacting with the endosomal SNARE syntaxin 13. We propose that GRASP-1 connects early and late recycling endosomal compartments by forming a molecular bridge between Rab-specific membrane domains and the endosomal SNARE machinery. The data uncover a new mechanism to achieve specificity and directionality in neuronal membrane receptor trafficking